Chicken embryo cultures in the dorsal-upward orientation for the manipulation of epiblasts

Chicken embryos have many advantages in the study of amniote embryonic development. In particular, culture techniques developed for early-stage embryos have promoted the advancement of modern developmental studies using chicken embryos. However, the standard technique involves placing chicken embryos in the ventral-upward (ventral-up) orientation, limiting manipulation of the epiblast at the dorsal surface, which is the primary source of ectodermal and mesodermal tissues. To circumvent this limitation, we developed chicken embryo cultures in the dorsal-up orientation and exploited this technique to address diverse issues. In this article, we first review the history of chicken embryo culture techniques to evaluate the advantages and limitations of the current standard technique. Then, the dorsal-up technique is discussed. These technological discussions are followed by three different examples of experimental analyses using dorsal-up cultures to illustrate their advantages: (1) EdU labeling of epiblast cells to assess potential variation in the cell proliferation rate; (2) migration behaviors of N1 enhancer-active epiblast cells revealed by tracking cells with focal fluorescent dye labeling in dorsal-up embryo culture; and (3) neural crest development of mouse neural stem cells in chicken embryos.